Phylogeny of Morella rubra and Its Relatives (Myricaceae) and Genetic Resources of Chinese Bayberry Using RAD Sequencing.

Liu L, Jin X, Chen N, Li X, Li P, Fu C - PLoS ONE (2015)

Bottom Line:
Here, we use restriction site-associated DNA sequencing (RAD-seq) to identify candidate loci that will help in determining phylogenetic relationships among Morella rubra, M. adenophora, M. nana and M. esculenta.Intraspecific divergence in M. rubra occurred in the late Pliocene (3.39 Ma).Additionally, we identified 3808 unlinked SNP sites across the four populations of M. rubra and discovered genes associated with fruit ripening and senescence, fruit quality and disease/defense metabolism based on KEGG database.

Affiliation: Key Laboratory of Conservation Biology for Endangered Wildlife of the Ministry of Education, and Laboratory of Systematic & Evolutionary Botany and Biodiversity, College of Life Sciences, Zhejiang University, Hangzhou 310058, China.

ABSTRACTPhylogenetic relationships among Chinese species of Morella (Myricaceae) are unresolved. Here, we use restriction site-associated DNA sequencing (RAD-seq) to identify candidate loci that will help in determining phylogenetic relationships among Morella rubra, M. adenophora, M. nana and M. esculenta. Three methods for inferring phylogeny, maximum parsimony (MP), maximum likelihood (ML) and Bayesian concordance, were applied to data sets including as many as 4253 RAD loci with 8360 parsimony informative variable sites. All three methods significantly favored the topology of (((M. rubra, M. adenophora), M. nana), M. esculenta). Two species from North America (M. cerifera and M. pensylvanica) were placed as sister to the four Chinese species. According to BEAST analysis, we deduced speciation of M. rubra to be at about the Miocene-Pliocene boundary (5.28 Ma). Intraspecific divergence in M. rubra occurred in the late Pliocene (3.39 Ma). From pooled data, we assembled 29378, 21902 and 23552 de novo contigs with an average length of 229, 234 and 234 bp for M. rubra, M. nana and M. esculenta respectively. The contigs were used to investigate functional classification of RAD tags in a BLASTX search. Additionally, we identified 3808 unlinked SNP sites across the four populations of M. rubra and discovered genes associated with fruit ripening and senescence, fruit quality and disease/defense metabolism based on KEGG database.

Mentions:
The single end of the paired end reads (R1) was applied for Phylogeny inference and divergence time estimation. We recovered c. 0.95×106 reads from each sample of Illumina sequencing. After filtering and clustering at 92% similarity with coverage greater than 6, we obtained c. 64,129 clusters per sample with a mean depth of 10.44. Around 33631 consensus loci passed filtering for paralogs (Table 3). The sequencing error (E = 1.63×10−3) was lower than heterozygosity (H = 8.18×10−3) by ML estimation. After clustering of consensus sequences across all 19 samples under the minimum taxa data set, we recovered 4253 loci including 8360 parsimony informative variable sites with 3677 unlinked SNP sites, which were applied for phylogeny inference of four species of Morella in China. For the Bayesian analysis, phylogeny reconstruction using the minimum taxa data set revealed strong support for the four species as a clade (1.00 PP), as well as the monophyly of each species. The topology of (((M. rubra, M. adenophora), M. nana), M. esculenta) was significantly favored. Two species from North America (M. cerifera and M. pensylvanica) were sister to the four Chinese species. The tree topologies from the MP and ML analyses (S1 Fig) were consistent with the results of the Bayesian analysis (Fig 2).

Mentions:
The single end of the paired end reads (R1) was applied for Phylogeny inference and divergence time estimation. We recovered c. 0.95×106 reads from each sample of Illumina sequencing. After filtering and clustering at 92% similarity with coverage greater than 6, we obtained c. 64,129 clusters per sample with a mean depth of 10.44. Around 33631 consensus loci passed filtering for paralogs (Table 3). The sequencing error (E = 1.63×10−3) was lower than heterozygosity (H = 8.18×10−3) by ML estimation. After clustering of consensus sequences across all 19 samples under the minimum taxa data set, we recovered 4253 loci including 8360 parsimony informative variable sites with 3677 unlinked SNP sites, which were applied for phylogeny inference of four species of Morella in China. For the Bayesian analysis, phylogeny reconstruction using the minimum taxa data set revealed strong support for the four species as a clade (1.00 PP), as well as the monophyly of each species. The topology of (((M. rubra, M. adenophora), M. nana), M. esculenta) was significantly favored. Two species from North America (M. cerifera and M. pensylvanica) were sister to the four Chinese species. The tree topologies from the MP and ML analyses (S1 Fig) were consistent with the results of the Bayesian analysis (Fig 2).

Bottom Line:
Here, we use restriction site-associated DNA sequencing (RAD-seq) to identify candidate loci that will help in determining phylogenetic relationships among Morella rubra, M. adenophora, M. nana and M. esculenta.Intraspecific divergence in M. rubra occurred in the late Pliocene (3.39 Ma).Additionally, we identified 3808 unlinked SNP sites across the four populations of M. rubra and discovered genes associated with fruit ripening and senescence, fruit quality and disease/defense metabolism based on KEGG database.

Affiliation:
Key Laboratory of Conservation Biology for Endangered Wildlife of the Ministry of Education, and Laboratory of Systematic & Evolutionary Botany and Biodiversity, College of Life Sciences, Zhejiang University, Hangzhou 310058, China.

ABSTRACTPhylogenetic relationships among Chinese species of Morella (Myricaceae) are unresolved. Here, we use restriction site-associated DNA sequencing (RAD-seq) to identify candidate loci that will help in determining phylogenetic relationships among Morella rubra, M. adenophora, M. nana and M. esculenta. Three methods for inferring phylogeny, maximum parsimony (MP), maximum likelihood (ML) and Bayesian concordance, were applied to data sets including as many as 4253 RAD loci with 8360 parsimony informative variable sites. All three methods significantly favored the topology of (((M. rubra, M. adenophora), M. nana), M. esculenta). Two species from North America (M. cerifera and M. pensylvanica) were placed as sister to the four Chinese species. According to BEAST analysis, we deduced speciation of M. rubra to be at about the Miocene-Pliocene boundary (5.28 Ma). Intraspecific divergence in M. rubra occurred in the late Pliocene (3.39 Ma). From pooled data, we assembled 29378, 21902 and 23552 de novo contigs with an average length of 229, 234 and 234 bp for M. rubra, M. nana and M. esculenta respectively. The contigs were used to investigate functional classification of RAD tags in a BLASTX search. Additionally, we identified 3808 unlinked SNP sites across the four populations of M. rubra and discovered genes associated with fruit ripening and senescence, fruit quality and disease/defense metabolism based on KEGG database.